A major focus of our work involves an evaluation of the acute and chronic effects of ethanol in the CNS. However, the brain represents a heterogeneous collection of cell types, and distinction of direct and indirect effects of ethanol can be difficult. In vitro cell culture systems can be used to monitor specific, direct effects of ethanol, for comparison and contrast with results obtained in brain ,tissue and in vivo. Using a primary culture of cerebellar granule cells, we found that ethanol, at low concentrations, inhibited glutamate (acting at kainate or NMDA receptors)-stimulated cyclic GMP production. Cyclic GMP production stimulated by atrial natriuretic peptide (ANP) was much less sensitive to ,inhibition by ethanol. The results suggested that glutamate receptor-effector coupling is altered by ethanol. Glutamate stimulates soluble guanylate cyclase in a calcium- dependent manner and preliminary findings indicate that ethanol alters glutamate-dependent calcium flux. These results suggest a new hypothesis for ethanol's depressant effects, and interaction of ethanol with NMDA-coupled calcium channels may also contribute to development of ethanol tolerance and dependence. In PC12 cells, we found that although ethanol stimulated adenylate cyclase (AC) activity in cell membranes, as in brain, ethanol inhibited agonist- stimulated cyclic AMP accumulation in one subclone of intact PC12 cells. In a second subclone ethanol often increased cyclic AMP levels. The inhibition of cyclic AMP production in the first subclone did not depend on Gi or on protein kinase C activity. The results demonstrate that the effect of ethanol on cyclic AMP accumulation depends on the characteristics of AC regulation in various cell types, so that extrapolations to other systems must be cautious. Nevertheless, the use of cell cultures allows investigations that can enhance our understanding of the sites and mechanisms of action of ethanol and of changes in biochemical systems that may, in vivo, be associated with ethanol tolerance and physical dependence.